Comparative genomics studies reveal the existence of long conserved noncoding sequences (CNSs) that are thought to play regulatory roles in the expression of mammalian genomes (Loots et al., Science, 288: 136-40 (2000); Dermitzakis et al., Nature, 420: 578-82 (2002)). Each type of nucleated cell of a mammal employs only a subset of the regulatory elements found in CNSs, thereby expressing only a subset of the genes of the mammalian cell at a given time. Many diseases and disorders are associated with abnormal gene expression, which can manifest as shifts in an affected cell's gene expression profile. Genetic modifications in a regulatory region of a CNS, i.e., changes in the regulatory sequence itself, can alter expression of an operably linked coding sequence. In addition, though all of the nucleated cells of a mammalian species have the same genome, every cell type has a different epigenome defined by posttranslational modifications of chromatin. Like changes to the actual regulatory sequence, post-translational modification of chromatin structure also can alter gene expression by, for example, allowing cellular proteins, such as transcription factors, access to DNA for transcription.
DNA is held in a chromatin structure, in part, by interactions with histone proteins. Histone modifications regulate the accessibility of chromatin and gene activity (reviewed in, for example, Kurdistani & Grunstein, Nat. Rev. Mol. Cell. Biol., 4: 276-84 (2003); Berger, Curr. Opin. Genet. Dev., 12: 142-48 (2002)). Histone acetylation is required for gene activation and cell growth (Durrin et al., Cell, 65: 1023-31 (1991); Megee et al., Science, 247: 841-45 (1990)). The mechanisms by which histone acetylation regulates chromatin structure and transcription are not fully understood; however, it is believed that the acetylation status of histone proteins signal factors that further regulate chromatin structure and gene activity (Strahl & Allis, Nature, 403: 41-45 (2000)). For example, the recruitment of Sir3 to form heterochromatin in yeast requires that the lysine at position 16 of the histone H4 protein is deacetylated (Hecht et al., Nature, 383: 92-96 (1996)). Therefore, acetylated histones may recruit and/or stabilize transcription factors and chromatin remodeling enzymes to their respective target sites in chromatin (Agalioti et al., Cell, 111: 381-92 (2002); Hassan et al., Cell, 104: 817-27 (2001)).
Delineating patterns of protein-DNA interactions and post-translational modifications to chromatin subunits would provide insight into the genomic positions of regulatory regions, the sequence of events leading to shifts in gene expression, and the type of protein-DNA interactions required for transcription. Currently, protein-DNA interactions are detected using marker and reporter proteins, which create a detectable signal when a desired protein binds its target sequence. Reporter assays require manipulation of the cellular environment. Thus, the results may not be predictive of the extent to which target proteins bind to target sequences in the absence of manipulation. Like reporter assays, electrophoretic mobility shift assays (EMSAs) provide little, if any, information regarding the genomic location of target sequences. Current methods also are time consuming and unspecific. DNA-protein interactions can be evaluated using chromatin immunoprecipitation (ChIP) in conjunction with DNA microarrays. However, currently available DNA microarrays generally cover only a small portion of a genome, which severely hinders analysis. In view of the above, there is a need in the art for an alternative method for identifying intracellular protein-DNA interactions. Ideally, the method can be extended to epigenomic research to analyze patterns of intracellular protein-DNA interactions associated with shifts in gene expression. The invention provides such a method. These and other advantages of the invention, as well as additional inventive features, will be apparent from the description of the invention provided herein.